Antibodies to double stranded (ds) DNA are not only highly specific for systemic lupus erythematosus (SLE), but are also directly involved in the pathogenesis of lupus nephritis (LN), a major disease manifestation. However, despite the clear association between anti-dsDNA antibodies and nephritis, the mechanisms by which anti-DNA antibodies contribute to renal damage have yet to be conclusively determined. LN may be initiated by formation of immune complexes in situ, via binding of anti-DNA antibodies to nucleosomal antigens deposited on the glomerular basement membrane (GBM). Alternatively, anti-DNA antibodies may be pathogenic not by virtue of binding to nuclear antigens, but rather via binding to cross reactive renal targets. In the initial period of funding for this grant, we found that pathogenic murine anti-DNA antibodies bind to mesangial cell (MC) (-actinin, and that high titers of anti-(-actinin antibodies were present in the serum and kidney eluates of lupus mice. Furthermore, we reported that in human SLE serum anti-(-actinin antibodies that bound to MC were present in high titers as well, and were closely associated with the presence and activity of LN. Finally, pathogenic antibodies binding to MC (-actinin directly modulated inflammatory gene expression including cytokines and neutrophil gelatinase associated lipocalin (NGAL, lipocalin-2), via Fc-dependent and independent mechanisms. We hypothesize that gene regulation induced by nephritogenic antibodies in kidney cells is an important contributor to the pathogenesis of LN, mediated by binding to cell surface receptors and engagement of an additional receptor from the Toll-like receptor (TLR) family, and that serum and/or urinary levels of NGAL may reflect the degree of renal injury induced by nephritogenic antibodies. We propose to continue our studies to understand the renal pathogenicity of anti-DNA antibodies. Specifically we will: I) Study the importance of specificity for (-actinin in determining antibody nephritogenicity and the effects of binding by pathogenic antibodies on (-actinin structure and cellular function; II) Investigate the mechanisms of direct gene modulation in kidney cells, including Fc receptor and TLR signaling pathways;and III) Determine if NGAL is a reliable marker for injury of kidney cells by nephritogenic antibodies, is NGAL upregulation instrumental in the pathogenesis of SLE renal disease, and whether serum and/or urine levels of NGAL may be useful as a biomarker for lupus nephritis. Public Health Relevance: Inflammation of the kidney, or nephritis, is a common and dangerous manifestation of disease in patients with systemic lupus erythematosus. While it is known that in lupus patients antibodies against DNA contribute to nephritis, how they actually cause kidney damage is not yet clear. We propose to study how anti-DNA antibodies induce kidney injury in an attempt to identify new approaches for more effective treatment, and discover novel serum or urine diagnostic markers that will improve the clinical care of lupus patients.